2.1. PLANT ELONGATION AND GROWTH
The plant cell elongation mechanism is a fundamental process with primary importance in plant-tissue development. Cell elongation requires relaxation of the rigid primary cell wall (Carpita and Gibeaut, 1993, Plant J. 3:1-30; Cosgrove, 1993, Plant Physiol. 102:1-6; Fry, 1988, The Growing Plant Cell Wall Chemical and Metabolic Analysis, Lonoman Scientific & Technical, New York; Roberts, 1994, Curr. Opin. Cell Biol. 6:688-694). Several mechanisms for this relaxation have been suggested, including the activities of endo-xyloglucan transferase (Nishitani and Tominaga, 1992, J. Biol. Chem. 267:21058-21064), xyloglucan endotransglycosylase (Fry et al., 1992, Biochem. J. 282:821-828) and expansins (McQueen-Mason and Cosgrove, 1995, Plant Physiol. 107:87-100). Endo-1,4-.beta.-glucanase (hereinafter, EGase) has been suggested to play an important role in the elongation process (Shoseyov and Dekel-Reichenbach, 1992, Acta Hort. 329:225-227; Verma et al., 1975, J. Biol. Chem.250:1019-1026).
Substantial evidence for the involvement of a 1,3-1,4-.beta.-glucan-specific enzyme in cell elongation was found in monocotyledons (Hatfield and Nevins, 1987, Plant Physiol. 83:203-207; Hoson and Nevins, 1989, Plant Physiol. 90:1353-1358 1989; Inouhe and Nevins, 1991, Plant Physiol. 96:426-431). EGase has been implicated in xyloglucan degradation during vegetative growth and fruit ripening (Hayashi, 1989, Ann. Rev. Plant Physiol. 40:139-168; Hayashi et al., 1984, Plant Physiol. 25:605-610). The activity of this enzyme could affect the generation of oligosaccharins, signaling molecules that are involved, among other things, in plant development and cell elongation (see for review, Darvill et al., 1992, Glycobiology 2:181-198).
To date, most of the EGase genes isolated have been studied in relation to fruit ripening (Cass et al., 1990, Mol. Gen. Genet. 223:76-86; Fischer and Bennett, 1991, Ann. Rev. Plant Physiol. Plant Mol. Biol. 42:675-703; Lashbrook et al., 1994, Plant Cell 6:1485-1493; Tucker et al., 1987, Plant Mol. Biol. 9:197-203) and abscission zones (Kemmerer and Tucker, 1994, Plant Physiol. 104:557-562; Tucker and Milligan, 1991, Plant Physiol. 95:928-933; Tucker et al., 1988, Plant Physiol. 88:1257-1262).
More recently, Wu et al. (1996, Plant Physiol. 110:163-170) cloned the EGase gene from pea and showed its expression to be induced by auxin in elongating epicotyls.
Endogenous regulation of cell elongation appears to be dominated by cell wall mechanics. This process is a result of the interaction between internal turgor pressure and the mechanical strength of the cell wall (reviewed by Steer and Steer, 1989, New Phytol. 111:323-358). Unlike most plant cells, the growth of pollen tubes and root hairs is restricted to the tip zone (reviewed by Cresti and Tiezzi, 1992, "Pollen tube emission organization and tip growth," in Sexual Plant Reproduction, pp. 89-97, eds. Cresti and Tiezzi, Springer-Verlag, Berlin). The growing region of pollen tubes consists of two distinct layers when fully mature. The inner layer consists mostly of callose-related molecules and the outer layer contains pectin, xyloglucan (XG), cellulose (at low levels and poor crystallinity) and other polysaccharides (reviewed by Steer and Steer, 1989, New Phytol. 111:323-358).
Xyloglucans (XGs) are linear chains of .beta.-(1-4)-D-glucan, but unlike cellulose, they possess numerous xylosyl units added at regular sites to the 0-6 position of the glucosyl units of the chain (reviewed by Carpita and Gibeaut, 1993, Plant J. 3:1-30). XG can be extracted by alkaline treatment and then bound again in vitro to cellulose (Hayashi et al., 1994, Plant Cell Physiol. 35:1199-1205).
XG is bound to cellulose microfibrils in the cell walls of all dicotyledons and some monocotyledons (reviewed by Roberts, 1994, Curr. Opin. Cell Biol. 6:688-694). The XG bound to the cellulose microfibrils cross-links the cell-wall framework.
Plant-cell expansion, including elongation, requires the integration of local wall-loosening and the controlled deposition of new wall materials. Fry et al. (1992, Biochem J. 282:821-828) and Nishitani and Tominaga (1992, J. Biol. Chem 267:21058-21064) purified xyloglucan endo-transglycosylase (XET) and endo-xyloglucan transferase (EXT), respectively. These two enzymes were shown to be responsible for the transfer of intermicrofibrillar XG from one segment to another XG molecule and thus, suggested to be wall loosening-enzymes.
However, McQueen-Mason et al. (1993, Planta 190:327-331) showed that XET activity did not correlate with in vitro cell wall extension in cucumber hypocotyls.
The effect of XG on growing tissues has been extensively investigated. XG oligosaccharides, produced by partial digestion with .beta.-(1-4)-D-glucanase and referred to as "oligosaccharins", alter plant-cell growth (reviewed by Aldington and Fry, 1993, Advances in Botanical Research 19:1-101). One such oligosaccharin, XXFG (XG9), antagonizes the growth promotion induced in pea stem segments by the auxin 2,4-D at a concentration of about 1 nM (York et al., 1984, Plant Physiol. 75:295-297; McDougall and Fry, 1988, Planta 175:412-416). On the other hand, at high concentrations (e.g., 100 .mu.M) oligosaccharins promote the elongation of etiolated pea stem segments (McDougall and Fry, 1990, Plant Physiol. 93:1042-1048). The mode of action of oligosaccharins is still unknown.
Another type of cell wall-loosening protein, termed "expansin", was isolated by McQueen-Mason et al. (1992, The Plant Cell 4:1425-1433). Expansin does not exhibit hydrolytic activity with any of the cell-wall components. It binds at the interface between cellulose microfibrils and matrix polysaccharides in the cell wall, and is suggested to induce cell wall expansion by reversibly disrupting noncovalent bonds within this polymeric network (McQueen-Mason and Cosgrove, 1995, Plant Physiol. 107:87-100). Some cellulose-binding organic substances alter cell growth and cellulose-microfibril assembly in vivo. Direct dyes, carboxymethyl cellulose (CMC) and fluorescent brightening agents (FBAs, e.g., calcofluor white ST) prevent Acetobacter xylinum microfibril crystallization, thereby enhancing polymerization. These molecules bind to the polysaccharide chains immediately after their extrusion from the cell surface, preventing normal assembly of microfibrils and cell walls (Haigler, 1991, "Relationship between polymerization and crystallization in microfibril biogenesis," in Biosynthesis and Biodegradation of Cellulose, pp. 99-124, Haigler and Weimer eds., Marcel Dekker, Inc., New York). Haigler discusses dyes and fluorescent brightening agents that bind to cellulose alter cellulose microfibril assembly in vivo. Modifications in cell shape were observed when red alga (Waaland and Waaland, 1975, Planta 126:127-138) and root tips (Hughes and McCully, 1975, Stain Technology 50:319-329) were grown in the presence of dyes. It is now evident that these molecules can bind to the cellulose chains immediately upon their extrusion from the cell surface of prokaryotes and eukaryotes (Haigler and Brown, 1979 Science 210:903-906; Benziman et al., 1980, Proc. Natl. Acad. Sci. USA 77:6678-6682; Haigler et al., 1980, Science 210:903-906; Brown et al., 1982, Science 218:1141-1142) and prevent crystal-structure formation (Haigler and Chanzy, 1988, J. Ultrastruct. Mol. Struct. Res. 98:299-311). In addition, the rate of cellulose polymerization was shown to increase in the presence of dye (Benziman et al., 1980). Crystallization was proposed to be the bottleneck in this coupled reaction and its prevention to result in accelerated cellulose synthase activity.